Fluorescent probes have been widely used in cell detection and measurement systems. Background fluorescence limits the ability of these systems to distinguish labeled cells from non-specific fluorescent signals. Background fluorescence can be generated when excess fluorescent probes exist outside the labeled cells. The excess probes can be freely dissolved in the test sample, producing a continuous, high level of background noise. They can also aggregate or bind to particulate materials in the test sample.
Unlike the freely dissolved probes, the fluorescent aggregates or particulates can be recognized as cells by detection instruments, leading to false positive signals. If there are large numbers of such particles, the fact that their fluorescent bursts occur very close together and at a high rate can overwhelm the data system of typical cell detection systems. Therefore, it is desirable to reduce background fluorescence produced by these aggregates and particulates.
U.S. Pat. No. 6,221,612 describes methods of using various dyes to reduce undesirable light emission. According to the disclosure, dyes which have absorption spectra that overlap with the absorption, emission or excitation spectrum of a fluorescent probe can be used to decrease background fluorescence generated by that probe. However, different dyes have different chemical and fluorescence properties. Selecting a suitable dye for a particular fluorescent probe and finding the proper conditions to use the dye may require extensive research.